人参皂苷Rh_2对食管癌细胞Eca-109细胞周期的影响

Effects of ginsenoside Rh_2(GS-Rh_2) on cell cycle of Eca-109 esophageal carcinoma cell line

目的:探讨人参皂苷Rh2(ginenoside-Rh2,GS-Rh2)对人食管癌细胞Eca-109生长抑制及细胞周期的影响。方法:应用MTT法测定GS-Rh2对细胞的生长抑制作用,HE染色后光镜观察细胞形态变化,应用免疫细胞化学和Western blot检测细胞周期调控因子cyclinE,CDK2(cyclin-dependent kinase 2),p21WAF1基因蛋白表达,采用半定量RT-PCR检测基因mRNA表达。结果:GS-Rh2对Eca-109细胞生长有抑制作用,并呈剂量、时间依赖性。20μg.mL-1GS-Rh2作用1 d后细胞达到半数抑制,作用3 d的Eca-109细胞较对照细胞数目明显减少,细胞形态向正常细胞方向逆转。细胞周期分析显示,随着GS-Rh2药物剂量的增加,G0/G1期细胞数逐渐增加,S和G2/M期细胞数逐渐减少,10μg.mL-1和20μg.mL-1GS-Rh2处理组和对照组比较有显著性差异。20μg.mL-1GS-Rh2处理细胞后,随着作用时间延长,cyclinE和CDK2蛋白及其mRNA表达水平逐渐增高,而p21WAF1蛋白及其mRNA表达水平逐渐降低。结论:GS-Rh2可以将人食管癌细胞Eca-109阻滞于G0/G1期,诱导肿瘤细胞分化,并且通过影响细胞周期调控因子cy-clinE,CDK2,p21WAF1基因的表达抑制食管癌细胞的增殖。

Objective： To investigate the effects of ginsenoside Rh2 （GS-Rh2） on growth inhibition and cell cycle of Eca-109 esophageal carcinoma cell line in culture. Method： The effects of GS-Rh2 on cell growth inhibition was detected by MTF assay. Cell cycle was analyzed by flow cytometry （FCM）. Cell morphology was observed by a light microscope after HE staining. The protein expression of cell cycle components （cyclinE, CDK2, p21^WAFI） were examined by immunocytochemistry and Western blot. The mRNA expression were examined by semiquantitative RT-PCR. Result： GS-Rh2 inhibited the proliferation of Eca-109 cells in dose and time-dependent manners. The inhibition rate was about 50% after 1-day treatment with 20μg·mL^-1 GS-Rh2,20μg·mL^-1 GS-Rh2 induced the mature differentiation and morphological reversion. With increasing dose of GS-Rh2 treatment, the cell number of G0/G1 phase was increased, whereas it decreased at S and G2/M phase. There was significant difference between 10, 20μg·mL^-1 GS-Rh2 groups and the corresponding group without GS-Rh2 treatement. After treating cells by20μg·mL^-1 GS-Rh2 for 1, 2, 3 days individually, the protein and mRNA expression of both cyclinE and CDK2 reduced ,while the expression of p21wan enhanced gradually. Conclusion： GS-Rh2 could arrest Eca-109 cells at G0/G1 phase and induce cell differentiation tending to normal. Furthermore, GS-Rh2 had an effect on expression of cell cycle components （cychnE , CDK2 and p21^WAFI ） to inhibit Eca-109 cell proliferation.